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Zeitschriftenartikel zum Thema "Recombinase RecA"
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del Val, Elsa, William Nasser, Hafid Abaibou und Sylvie Reverchon. „RecA and DNA recombination: a review of molecular mechanisms“. Biochemical Society Transactions 47, Nr. 5 (18.10.2019): 1511–31. http://dx.doi.org/10.1042/bst20190558.
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Abstract Recombinases are responsible for homologous recombination and maintenance of genome integrity. In Escherichia coli, the recombinase RecA forms a nucleoprotein filament with the ssDNA present at a DNA break and searches for a homologous dsDNA to use as a template for break repair. During the first step of this process, the ssDNA is bound to RecA and stretched into a Watson–Crick base-paired triplet conformation. The RecA nucleoprotein filament also contains ATP and Mg2+, two cofactors required for RecA activity. Then, the complex starts a homology search by interacting with and stretching dsDNA. Thanks to supercoiling, intersegment sampling and RecA clustering, a genome-wide homology search takes place at a relevant metabolic timescale. When a region of homology 8–20 base pairs in length is found and stabilized, DNA strand exchange proceeds, forming a heteroduplex complex that is resolved through a combination of DNA synthesis, ligation and resolution. RecA activities can take place without ATP hydrolysis, but this latter activity is necessary to improve and accelerate the process. Protein flexibility and monomer–monomer interactions are fundamental for RecA activity, which functions cooperatively. A structure/function relationship analysis suggests that the recombinogenic activity can be improved and that recombinases have an inherently large recombination potential. Understanding this relationship is essential for designing RecA derivatives with enhanced activity for biotechnology applications. For example, this protein is a major actor in the recombinase polymerase isothermal amplification (RPA) used in point-of-care diagnostics.
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Luisi-DeLuca, C., S. T. Lovett und R. D. Kolodner. „Genetic and physical analysis of plasmid recombination in recB recC sbcB and recB recC sbcA Escherichia coli K-12 mutants.“ Genetics 122, Nr. 2 (01.06.1989): 269–78. http://dx.doi.org/10.1093/genetics/122.2.269.
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Abstract The effect of mutations in known recombination genes (recA, recB, recC, recE, recF, recJ, recN, recO, recQ and ruv) on intramolecular recombination of plasmids was studied in recB recC sbcB and recB recC sbcA Escherichia coli mutants. The rate of recombination of circular dimer plasmids was at least 1000-fold higher in recB recC sbcB or recB recC sbcA mutants as compared to wild-type cells. The rate was decreased by mutations in recA, recF, recJ, recO, ruv or mutS in recB recC sbcB mutants, and by mutations in recE, recN, recO, recQ, ruv or mutS in recB recC sbcA mutants. In addition to measuring the recombination rate of circular dimer plasmids, the recombination-mediated transformation of linear dimer plasmids was also studied. Linear dimer plasmids transformed recB recC sbcB and recB recC sbcA mutants 20- to 40-fold more efficiently than wild-type cells. The transformation efficiency of linear dimer plasmids in recB recC sbcB mutants was decreased by mutations in recA, recF, recJ, recO, recQ or lexA (lexA3). In recB recC sbcA mutants the transformation efficiency of linear dimers was decreased only by a recE mutation. Physical analysis of linear dimer- or circular dimer-transformed recB recC sbcB mutants revealed that all transformants contained recombinant monomer genotypes. This suggests that recombination in recB recC sbcB cells is very efficient.
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Chittela, Rajani Kant, und Jayashree K. Sainis. „Plant DNA Recombinases: A Long Way to Go“. Journal of Nucleic Acids 2010 (2010): 1–10. http://dx.doi.org/10.4061/2010/646109.
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DNA homologous recombination is fundamental process by which two homologous DNA molecules exchange the genetic information for the generation of genetic diversity and maintain the genomic integrity. DNA recombinases, a special group of proteins bind to single stranded DNA (ssDNA) nonspecifically and search the double stranded DNA (dsDNA) molecule for a stretch of DNA that is homologous with the bound ssDNA. Recombinase A (RecA) has been well characterized at genetic, biochemical, as well as structural level from prokaryotes. Two homologues of RecA called Rad51 and Dmc1 have been detected in yeast and higher eukaryotes and are known to mediate the homologous recombination in eukaryotes. The biochemistry and mechanism of action of recombinase is important in understanding the process of homologous recombination. Even though considerable progress has been made in yeast and human recombinases, understanding of the plant recombination and recombinases is at nascent stage. Since crop plants are subjected to different breeding techniques, it is important to know the homologous recombination process. This paper focuses on the properties of eukaryotes recombinases and recent developments in the field of plant recombinases Dmc1 and Rad51.
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Haldenby, Sam, Malcolm F. White und Thorsten Allers. „RecA family proteins in archaea: RadA and its cousins“. Biochemical Society Transactions 37, Nr. 1 (20.01.2009): 102–7. http://dx.doi.org/10.1042/bst0370102.
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Recombinases of the RecA family are essential for homologous recombination and underpin genome stability, by promoting the repair of double-stranded DNA breaks and the rescue of collapsed DNA replication forks. Until now, our understanding of homologous recombination has relied on studies of bacterial and eukaryotic model organisms. Archaea provide new opportunities to study how recombination operates in a lineage distinct from bacteria and eukaryotes. In the present paper, we focus on RadA, the archaeal RecA family recombinase, and its homologues in archaea and other domains. On the basis of phylogenetic analysis, we propose that a family of archaeal proteins with a single RecA domain, which are currently annotated as KaiC, be renamed aRadC.
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García-Vázquez, Francisco A., Salvador Ruiz, Carmen Matás, M. José Izquierdo-Rico, Luis A. Grullón, Aitor De Ondiz, Luis Vieira, Karen Avilés-López, Alfonso Gutiérrez-Adán und Joaquín Gadea. „Production of transgenic piglets using ICSI–sperm-mediated gene transfer in combination with recombinase RecA“. REPRODUCTION 140, Nr. 2 (August 2010): 259–72. http://dx.doi.org/10.1530/rep-10-0129.
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Sperm-mediated gene transfer (SMGT) is a method for the production of transgenic animals based on the intrinsic ability of sperm cells to bind and internalize exogenous DNA molecules and to transfer them into the oocyte at fertilization. Recombinase-A (RecA) protein-coated exogenous DNA has been used previously in pronuclear injection systems increasing integration into goat and pig genomes. However, there are no data regarding transgene expression after ICSI. Here, we set out to investigate whether the expression of transgenic DNA in porcine embryos is improved by recombinase-mediated DNA transfer and if it is possible to generate transgenic animals using this methodology. Different factors which could affect the performance of this transgenic methodology were analyzed by studying 1) the effect of the presence of exogenous DNA and RecA protein on boar sperm functionality; 2) the effect of recombinase RecA on in vitro enhanced green fluorescent protein (EGFP)-expressing embryos produced by ICSI or IVF; and 3) the efficiency of generation of transgenic piglets by RecA-mediated ICSI. Our results suggested that 1) the presence of exogenous DNA and RecA–DNA complexes at 5 μg/ml did not affect sperm functionality in terms of motility, viability, membrane lipid disorder, or reactive oxygen species generation; 2) EGFP-expressing embryos were obtained with a high efficiency using the SMGT–ICSI technique in combination with recombinase; however, the use of IVF system did not result in any fluorescent embryos; and 3) transgenic piglets were produced by this methodology. To our knowledge, this is the first time that transgenic pigs have been produced by ICSI-SGMT and a recombinase.
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Hofstatter, Paulo G., Alexander K. Tice, Seungho Kang, Matthew W. Brown und Daniel J. G. Lahr. „Evolution of bacterial recombinase A ( recA ) in eukaryotes explained by addition of genomic data of key microbial lineages“. Proceedings of the Royal Society B: Biological Sciences 283, Nr. 1840 (12.10.2016): 20161453. http://dx.doi.org/10.1098/rspb.2016.1453.
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Recombinase enzymes promote DNA repair by homologous recombination. The genes that encode them are ancestral to life, occurring in all known dominions: viruses, Eubacteria, Archaea and Eukaryota. Bacterial recombinases are also present in viruses and eukaryotic groups (supergroups), presumably via ancestral events of lateral gene transfer. The eukaryotic recA genes have two distinct origins (mitochondrial and plastidial), whose acquisition by eukaryotes was possible via primary (bacteria–eukaryote) and/or secondary (eukaryote–eukaryote) endosymbiotic gene transfers (EGTs). Here we present a comprehensive phylogenetic analysis of the recA genealogy, with substantially increased taxonomic sampling in the bacteria, viruses, eukaryotes and a special focus on the key eukaryotic supergroup Amoebozoa, earlier represented only by Dictyostelium . We demonstrate that several major eukaryotic lineages have lost the bacterial recombinases (including Opisthokonta and Excavata), whereas others have retained them (Amoebozoa, Archaeplastida and the SAR-supergroups). When absent, the bacterial recA homologues may have been lost entirely (secondary loss of canonical mitochondria) or replaced by other eukaryotic recombinases. RecA proteins have a transit peptide for organellar import, where they act. The reconstruction of the RecA phylogeny with its EGT events presented here retells the intertwined evolutionary history of eukaryotes and bacteria, while further illuminating the events of endosymbiosis in eukaryotes by expanding the collection of widespread genes that provide insight to this deep history.
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Liu, Yu-Tien, Chia-Geun Chen, Der-Chiang Chao, Fan Lee, Ching-Len Liao, Huey-Kang Sytwu, Chi-Fu Chou und Dar-Der Ji. „Sequence analysis of theGluconobacter oxydansRecA protein and construction of arecA-deficient mutant“. Canadian Journal of Microbiology 45, Nr. 4 (01.04.1999): 347–51. http://dx.doi.org/10.1139/w99-009.
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The deduced amino acid sequence of Gluconobacter oxydans RecA protein shows 75.2, 69.4, and 66.2% homology with those from Aquaspirillum magnetotacticum, Escherichia coli, andPseudomonas aeruginosa, respectively. The amino acid residues essential for function of the recombinase, protease, and ATPase in E. coli recA protein are conserved in G. oxydans. Of 24 amino acid residues believed to be the ATP binding domain of E. coli RecA, 17 are found to be identical in G. oxydans RecA. Interestingly, nucleotide sequence alignment between the SOS box of G. orphans recA gene and those from different microorganisms revealed that all the DNA sequences examined have dyad symmetry that can form a stem-loop structure. A G. oxydans recA-deficient mutant (LCC96) was created by allelic exchange using the cloned recA gene that had been insertionally inactivated by a kanamycin-resistance cassette. Such replacement of the wild-type recA with a kanamycin resistance gene in the chromosome was further verified by Southern hybridization. Phenotypically, the recA-deficient mutant is significantly more sensitive to UV irradiation than the wild-type strain, suggesting that the recA gene of G. oxydans ATCC9324 plays a role in repairing DNA damage caused by UV irradiation. Moreover, the mutant strain is much more plasmid transformable than its parent strain, illustrating that G. oxydans LCC96 could be used as a host to take up the recombinant plasmid for gene manipulation.Key words: Gluconobacter orphans, recA gene, DNA repair, recA mutant, SOS box.
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Inagaki, Satoko, Kazuyo Fujita, Yukiko Takashima, Kayoko Nagayama, Arifah C. Ardin, Yuki Matsumi und Michiyo Matsumoto-Nakano. „Regulation of Recombination betweengtfB/gtfCGenes inStreptococcus mutansby Recombinase A“. Scientific World Journal 2013 (2013): 1–7. http://dx.doi.org/10.1155/2013/405075.
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Streptococcus mutansproduces 3 types of glucosyltransferases (GTFs), whose cooperative action is essential for cellular adhesion. The recombinase A (RecA) protein is required for homologous recombination. In our previous study, we isolated several strains with a smooth colony morphology and low GTF activity, characteristics speculated to be derived from the GTF fusions. The purpose of the present study was to investigate the mechanism of those fusions.S. mutansstrain MT8148 was grown in the presence of recombinant RecA (rRecA) protein, after which smooth colonies were isolated. The biological functions and sequences of thegtfBandgtfCgenes of this as well as other clinical strains were determined. The sucrose-dependent adherence rates of those strains were reduced as compared to that of MT8148. Determination of the sequences of thegtfBandgtfCgenes showed that an approximately 3500 bp region was deleted from the area between them. Furthermore, expression of therecAgene was elevated in those strains as compared to MT8148. These results suggest that RecA has an important role in fusions ofgtfBandgtfCgenes, leading to alteration of colony morphology and reduction in sucrose-dependent adhesion.
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Pan, Yue, Ningkang Xie, Xin Zhang, Shuo Yang und Shaowu Lv. „Computational Insights into the Dynamic Structural Features and Binding Characteristics of Recombinase UvsX Compared with RecA“. Molecules 28, Nr. 8 (11.04.2023): 3363. http://dx.doi.org/10.3390/molecules28083363.
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RecA family recombinases are the core enzymes in the process of homologous recombination, and their normal operation ensures the stability of the genome and the healthy development of organisms. The UvsX protein from bacteriophage T4 is a member of the RecA family recombinases and plays a central role in T4 phage DNA repair and replication, which provides an important model for the biochemistry and genetics of DNA metabolism. UvsX shares a high degree of structural similarity and function with RecA, which is the most deeply studied member of the RecA family. However, the detailed molecular mechanism of UvsX has not been resolved. In this study, a comprehensive all-atom molecular dynamics simulation of the UvsX protein dimer complex was carried out in order to investigate the conformational and binding properties of UvsX in combination with ATP and DNA, and the simulation of RecA was synchronized with the property comparison learning for UvsX. This study confirmed the highly conserved molecular structure characteristics and catalytic centers of RecA and UvsX, and also discovered differences in regional conformation, volatility and the ability to bind DNA between the two proteins at different temperatures, which would be helpful for the subsequent understanding and application of related recombinases.
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Ramos, Cristina, Rogelio Hernández-Tamayo, María López-Sanz, Begoña Carrasco, Ester Serrano, Juan C. Alonso, Peter L. Graumann und Silvia Ayora. „The RecD2 helicase balances RecA activities“. Nucleic Acids Research 50, Nr. 6 (02.03.2022): 3432–44. http://dx.doi.org/10.1093/nar/gkac131.
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Abstract DNA helicases of the RecD2 family are ubiquitous. Bacillus subtilis RecD2 in association with the single-stranded binding protein SsbA may contribute to replication fork progression, but its detailed action remains unknown. In this work, we explore the role of RecD2 during DNA replication and its interaction with the RecA recombinase. RecD2 inhibits replication restart, but this effect is not observed in the absence of SsbA. RecD2 slightly affects replication elongation. RecA inhibits leading and lagging strand synthesis, and RecD2, which physically interacts with RecA, counteracts this negative effect. In vivo results show that recD2 inactivation promotes RecA–ssDNA accumulation at low mitomycin C levels, and that RecA threads persist for a longer time after induction of DNA damage. In vitro, RecD2 modulates RecA-mediated DNA strand-exchange and catalyzes branch migration. These findings contribute to our understanding of how RecD2 may contribute to overcome a replicative stress, removing RecA from the ssDNA and, thus, it may act as a negative modulator of RecA filament growth.
Dissertationen zum Thema "Recombinase RecA"
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Mah, Wayne. „Single molecule study of RecA recombinase enzyme activity“. Thesis, McGill University, 2008. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=18743.
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Homologous recombination is an essential pathway in the repair of DNA damage during the DNA replication process. RecA protein promotes the central steps in homologous recombination, after coating single-stranded DNA (ssDNA), RecA carries out a pairing and strand exchange reaction involving homologous DNA. This research project aims to characterize RecA function in homologous recombination using single molecule tethered particle motion (TPM). Using TPM to observe RecA extension along DNA, the RecA extension rate on ssDNA was determined for the first time. The rate obtained for dsDNA was similar, implying that RecA polymerizes along only one strand of a DNA substrate. The nucleation behaviour of RecA on DNA was also obtained from the extension trace, confirming the hypothesis that rapid nucleation on ssDNA is pH independent, while nucleation on dsDNA is pH dependent. Several pilot single molecule experiments aimed at monitoring the pairing and strand exchange reaction in real time were attempted. Although these experiments were unsuccessful, successful ensemble biochemical analogues of these experiments proved the feasibility of the single molecule experiments. These attempts gave insights into possible factors hindering success and led to experimental suggestions essential to the success of future experiments.La recombinaison Homologue est un chemin essentiel dans la réparation de dommages d'ADN pendant le procédé de réplication d'ADN. La protéine de RecA promeut les étapes centrales dans la recombinaison homologue, après avoir revêtu ADN seul-abandonné (ssDNA), RecA exécute un mettre et la réaction d'échange de brin impliquant ADN homologue. Ce projet de recherche vise à caractériser la fonction de RecA dans la recombinaison homologue utilisant la molécule seule mouvement de particule attaché (TPM). TPM d'utilisation pour observer l'extension de RecA le long d'ADN, le taux d'extension de RecA sur ssDNA a été déterminé pour la première fois. Le taux obtenu pour dsDNA était similaire, impliquant ce RecA polymerizes le long de seulement un brin d'un substrat d'ADN. Le comportement de nucleation de RecA sur ADN a été aussi obtenu de la trace d'extension, confirmant l'hypothèse ce nucleation rapide sur ssDNA est indépendant du pH, pendant que nucleation sur dsDNA est dépendant du pH. Plusieurs pilote plusieurs expériences de molécule seules ont visé à contrôlant le mettre et la réaction d'échange de brin a été tentée en temps réel. Bien que ces expériences étaient les ensembles infructueuses et réussies analogues biochimiques de ces expériences ont prouvé la possibilité des expériences de molécule seules. Ces tentatives ont donné de l'aux perspicacités dans les facteurs possibles freinant le succès et a mené à l'élément essentiel de suggestions expérimental au succès d'expériences futures
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Perry, Thomas. „Étude structurale et fonctionnelle de l'appareil de recombinaison homologue chez Streptococcus pneumoniae“. Electronic Thesis or Diss., Sorbonne université, 2019. http://www.theses.fr/2019SORUS300.
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La transformation naturelle est la capacité de certaines bactéries à incorporer et à intégrer activement de l’ADN extra-cellulaire. Ce procédé majeur augmente la plasticité et l’adaptabilité des bactéries à Gram positif et négatif en réalisant des échanges génétiques intra- et inter-espèces. S. pneumoniae est un pathogène majeur de l’Homme et se retrouve de façon commensale dans les muqueuses du nasopharynx. Cette bactérie est responsable d’infections sévères telles que des pneumonies, des méningites et des septicémies. Dans cette espèce, la transformation naturelle est corrélée au phénomène de changement de capsule et à la baisse d’efficacité des vaccins, ainsi qu’à l’obtention de gènes de résistance aux antibiotiques. Au cours de la transformation, l’ADN extra-cellulaire va être pris en charge dans le cytoplasme par différentes protéines, qui permettront à terme d’intégrer ce gène au génome bactérien par recombinaison homologue. La recherche d’homologie et l’intégration du gène est sous le contrôle de la recombinase universelle RecA. Pour cela, RecA va former un filament hélicoïdal avec l’ADN sb exogène et db endogène, et ceux de manière ATP dépendante. Toutefois, la structure atomique de tels filaments n’a jamais été observé chez S. pneumoniae. Par des techniques de caractérisation biochimique et de cryo-microscopie électronique nous sommes parvenus à résoudre la structure des deux types de filaments à des résolutions de 3.8 Å et 3.9 Å. Ce qui nous a permis par la suite de mieux caractériser l’interaction de RecA avec l’ADN. En comparant nos structures avec celles obtenues par cristallographie des filaments de RecA observé chez E. coli, nous avons en partie expliqué les raisons de l’efficacité de recombinaison 3 fois supérieur de S. pneumoniae. Dans un second temps, nous avons mis en évidence in vitro un lien entre la filamentation de RecA le long de l’ADN et l’activité hélicase de RadA. RadA est une protéine nécessaire à la recombinaison homologue et dont l’activité hélicase semble promouvoir et étendre la D-loop au niveau de l’ADN db endogène. Nous avons également caractérisé, par RMN, la structure du motif en doigt de zinc de RadA et son interaction avec l’ADN, motif qui n’avait pu être résolu dans la structure de RadA en cristallographie et qui semble indispensable à son activité hélicaseNatural transformation is the ability of some bacteria to incorporate and actively integrate extracellular DNA. This major process increases the plasticity and adaptability of gram positive and negative bacteria by performing intra- and inter-species genetic exchanges. S. pneumoniae is a major human pathogen and is found commensally in the mucous membranes of the nasopharynx. This bacterium is responsible for severe infections such as pneumonia, meningitis and sepsis. In this species, the natural transformation is correlated with the phenomenon of capsule change and the decrease of vaccine efficacy, as well as the obtaining of genes for antibiotic resistance. During the transformation, the extracellular DNA will be supported in the cytoplasm by different proteins, which will eventually integrate this gene into the bacterial genome by homologous recombination. The search for homology and the integration of the gene is under the control of the universal recombinase RecA. For this, RecA will form a helical filament with exogenous ssDNA and endogenous dsDNA, and those in ATP dependent manner. However, the atomic structure of such filaments has never been observed in S. pneumoniae. Using biochemical characterization techniques and electron cryomicroscopy, we succeeded in solving the structure of the two types of filaments at resolutions of 3.8 Å and 3.9 Å. This allowed us to better characterize the interaction of RecA with DNA. By comparing our structures with those obtained by crystallography of RecA filaments observed in E. coli, we partly explained the reasons for the 3-fold recombination efficiency of S. pneumoniae. In a second step, we demonstrated in vitro a link between the RecA filamentation along the DNA and the helicase activity of RadA. RadA is a protein necessary for homologous recombination and whose helicase activity seems to promote and extend D-loop at the level of endogenous dsDNA. We have also characterized, by NMR, the structure of the zinc finger motif of RadA and its interaction with DNA, a motif that could not be solved in the RadA structure in crystallography and which seems essential for its helicase activity
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Kobir, Ahasanul. „Physiological roles of Eukaryotic Hanks type Ser/Thr kinase in transition to stationary phase in Bacillus subtilis“. Phd thesis, Université Paris Sud - Paris XI, 2012. http://tel.archives-ouvertes.fr/tel-00911812.
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Bacillus subtilis is the model organism for low GC Gram-positive bacteria and is of great biotechnological interest. Protein phosphorylation is an important regulatory mechanism in bacteria and it has not been extensively studied yet. Recent site-specific phosphoproteomic studies identified a large number of novel serine/threonine phosphorylation sites in B. subtilis, including a) two transition phase global gene regulators DegS and AbrB and b) RecA, that plays a major role in double-strand break repair and DNA recombination. .B. subtilis disposes of several putative Ser/Thr kinases like PrkA, YbdM, YabT and a characterizd kinase PrkC, but very few physiological substrates for these have been defined so far. In vitro phosphorylation assays were used to identify which of these kinases were able to phosphorylate DegS, RecA and AbrB. DegS phosphorylation on serine 76 by the kinase YbdM influenced its activity towards DegU both in vitro and in vivo, and expression of DegS S76D( on replacing serine to aspartate) in B. subtilis perturbed cellular processes regulated by the DegS/DegU two component system. This suggests a link between DegS phosphorylation at serine 76 and the level of DegU phosphorylation, establishing this post-translational modification as an additional trigger for this two-component system. At the onset of sporulation, B. subtilis expresses an unusual serine/threonine kinase YabT, which exhibits a septal localization and is activated by non-sequence-specific DNA binding. Activated YabT phosphorylates RecA at the residue serine 2, which in turn promotes the formation of RecA foci at the onset of spore development. On the other hand, non-phosphorylatable RecA or inactivated YabT lead to reduced spore formation in the presence of DNA lesions . This suggests a functional similarity between B. subtilis developmental stage dependent RecA phosphorylation and its eukaryal homologous Rad51 phosphorylation, which leads to its recruitment to the lesion sites. We therefore proposed that RecA phosphorylation serves as an additional signal mechanism that promotes focus formation during spore development. AbrB is phosphorylated by YabT, YbdM and PrkC in vitro and AbrB phosphorylation leads to reduced affinity for its target DNA and abolished binding cooperativity in vitro and in vivo. Expression of the phosphomimetic AbrB-S86D or of the non-phosphorylatable AbrB-S86A mutant protein in B. subtilis disturbed some stationary phase phenomena such as exoprotease production, competence and the onset of sporulation, probably by deregulation of AbrB-target genes and operons. We therefore, proposed that AbrB phosphorylation as an additional regulatory mechanism needed to switch off this ambiactive gene regulator during the transition phase.
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Dulermo, Rémi. „Etude des mécanismes de l'extrême tolérance aux radiations de la bactérie Deinococcus deserti par une approche de génomique fonctionnelle“. Aix-Marseille 2, 2009. http://theses.univ-amu.fr.lama.univ-amu.fr/2009AIX22100.pdf.
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Le génome de Deinococcus deserti, une bactérie très radiotolérante, a été analysé et comparé à ceux de D. Radiodurans et D. Geothermalis. Environ 230 protéines sont spécifiquement conservées chez ces 3 espèces, dont IrrE, un régulateur essentiel pour la radiotolérance. D. Deserti possède plusieurs gènes supplémentaires liés à la réparation de l’ADN, dont imuY et dnaE2 (ADN polymérases translesionnelles). En plus, D. Deserti a 3 recA qui codent pour 2 protéines RecA différentes (RecAC et RecAP). Pour étudier ces gènes, des outils génétiques ont été mis au point. Différents résultats suggèrent qu’IrrE, nécessaire pour l’induction de plusieurs gènes après irradiation, a une activité peptidase. Les 2 RecA sont fonctionnelles pour la réparation de l’ADN. D. Deserti est mutable par UV, ce qui nécessite ImuY, DnaE2 et RecAC, mais pas RecAPThe genome of Deinococcus deserti, a highly radiation-tolerant bacterium, was analyzed and compared to those of D. Radiodurans and D. Geothermalis. About 230 proteins are specifically conserved in these 3 species, including IrrE, a regulator protein essential for radiotolerance. D. Deserti has several supplementary DNA repair genes, like imuY and dnaE2 (translesion DNA polymerases). Moreover, D. Deserti has 3 recA that code for 2 different RecA proteins (RecAC et RecAP). To study these genes, genetic tools were developed for D. Deserti. Different results suggest that IrrE, required for the induction of several genes after irradiation, has peptidase activity. The 2 RecA proteins are functional for DNA repair. D. Deserti is mutable by UV, which requires ImuY, DnaE2 and RecAC, but not RecAP
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Chen, Zhucheng. „Mechanism of homologous recombination : from crystal structures of RecA-single stranded DNA and RecA-double stranded DNA filaments /“. Access full-text from WCMC, 2009. http://proquest.umi.com/pqdweb?did=1619205721&sid=8&Fmt=2&clientId=8424&RQT=309&VName=PQD.
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Ramdas, Jyoti. „Functions Of Nucleosomes And Other Regulatory Factor(S) In Homologous Recombination Promoted By RecA Protein“. Thesis, Indian Institute of Science, 1994. https://etd.iisc.ac.in/handle/2005/99.
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Homologous genetic recombination occurs during the life cycle of virtually every organism Genetic studies especially in prokaryotes and fungi have defined the rules of recombination, led to the characterization of alternate pathways and to the development of molecular models The biochemistry of homologous genetic recombination has advanced most productively in bacteria and fungi due to the extensive genetic understanding of these organisms The identification of mutants defective in homologous recombination, purification and characterization of the gene products that participate in recombination has brought the ultimate goal of reconstituting a cell-k free system for Eschenchia coli, at least with naked DNA substrates, closer to reality.
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Ramdas, Jyoti. „Functions Of Nucleosomes And Other Regulatory Factor(S) In Homologous Recombination Promoted By RecA Protein“. Thesis, Indian Institute of Science, 1994. http://hdl.handle.net/2005/99.
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Homologous genetic recombination occurs during the life cycle of virtually every organism Genetic studies especially in prokaryotes and fungi have defined the rules of recombination, led to the characterization of alternate pathways and to the development of molecular models The biochemistry of homologous genetic recombination has advanced most productively in bacteria and fungi due to the extensive genetic understanding of these organisms The identification of mutants defective in homologous recombination, purification and characterization of the gene products that participate in recombination has brought the ultimate goal of reconstituting a cell-k free system for Eschenchia coli, at least with naked DNA substrates, closer to reality.
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Saladin, Adrien. „Macromolecular Docking : applications to the RecA nucleofilament“. Paris 7, 2009. http://www.theses.fr/2009PA077098.
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Les protéines jouent un rôle central dans de nombreux processus cellulaire et peuvent intervenir dans de nombreuses interactions différentes, avec d'autres protéines, de l'ADN, des lipides, ou de petits ligands. La détermination de ces interactions est fondamentale pour pouvoir comprendre des processus biologiques majeurs et de nombreuses méthodes expérimentales ont été développées pour les caractériser. Cependant les méthodes expérimentales sont longues et coûteuses et les méthodes informatisées de prédictions d'interactions pourraient, à terme, fournir dans ce contexte une aide précieuse permettant de guider de futures expériences en biochimie et biologie moléculaire. Le développement logiciels d'amarrage est également un processus difficile mettant en jeu des cycles de conception d'algorithme, d'implémentation et de tests. Au cours de ma thèse, j'ai développé une librairie orientée objet pour favoriser et accélérer les étapes d'implémentation et de tests des méthodes d'amarrage. Cette librairie, programmée en C++ et interfacée avec le langage de script Python, a été utilisée pour mettre au point et tester de nouvelles méthodes appliquées à l'amarrage protéine-ADN et à l'amarrage multi-composants. Des programmes développés à l'aide de cette librairie sont actuellement appliqués à l'étude des modes d'amarrage de l'ADN au complexe RecA, responsable de la recombinaison homologue chez les bactériesProteins play a central role in various cellular processes with various interactions with other proteins, DNA, lipids or small ligands. Because the determination of these interactions is fundamental for understanding key biological processes, several experimental methods have been developed to characterize them. Experimental studies can take a long time and an expensive. Computational methods can therefore be of great help to guide future biochemical experiments. Development of docking software is a long process involving cycles of algorithm conception, programming and tests. During my thesis, I developed an object oriented library to help and speed-up development and tests of docking methods. This library was programmed in C++ with Python bindings, and has been used to test new methods applied to protein-DNA docking and multicomponent docking. Programs made with the help of this library are presently used to study the binding of DNA to the RecA complex, responsible of homologous recombination
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黃楚華 und Choi-wah Brian Wong. „In vitro studies on the mechanism of homologous DNA recombination promoted by Escherichia coli RecA protein“. Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 1993. http://hub.hku.hk/bib/B31233284.
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Wong, Choi-wah Brian. „In vitro studies on the mechanism of homologous DNA recombination promoted by Escherichia coli RecA protein /“. [Hong Kong : University of Hong Kong], 1993. http://sunzi.lib.hku.hk/hkuto/record.jsp?B13408902.
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Suzuki. Genetic Analysis, 4/E and Reco: Beyond the Myth of. W.H. Freeman & Company, 1992.
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Organisation for economic co-operation and development. Recombinant DNA Safety Considerations (Safety Considerations for Industrial, Agricultural and Environmental Applications of Organisms Derived By Reco). Organization for Economic, 1986.
Den vollen Inhalt der Quelle findenKonferenzberichte zum Thema "Recombinase RecA"
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Duarte, Gabriela Alves Carvalho, Vanessa Bridi, Dhullya Eduarda Resende Santos und Hanstter Hallison Alves Rezende. „TECNOLOGIA DO DNA RECOMBINANTE NA PRODUÇÃO DE INSULINA“. In I Congresso de Engenharia de Biotecnologia. Revista Multidisciplinar de Educação e Meio Ambiente, 2021. http://dx.doi.org/10.51189/rema/1377.
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Introdução: A diabetes mellitus (DM) é uma doença metabólica, que gera hiperglicemia. Decorre devido à uma deficiência nas células beta pancreáticas, responsável por produzir o hormônio da insulina, ou a uma resistência à insulina, sendo classificadas como DM tipo 1 e DM tipo 2. Tanto tipo 1, quanto tipo 2 severa, são tratadas com reposição de insulina. O avanço da engenharia genética, especialmente a tecnologia do DNA recombinante, possibilitou a criação de novas terapias gênicas, baseadas na programação de células, principalmente bacterianas, como Escherichia coli, utilizando fragmentos do plasmídeo bacteriano, recombinados à fragmentos do DNA humano, com a característica desejada. Ao introduzir esse DNA recombinado na célula hospedeira ocorrerá multiplicação do plasmídeo recombinante e divisão das bactérias. Objetivo: Descrever como a tecnologia do DNA recombinante foi aplicada na produção de insulina, e consequentemente no aprimoramento do tratamento de diabetes mellitus. Materiais e métodos: Refere-se à uma revisão bibliográfica, no qual foram utilizados os bancos de dados, PUBMED, SciELO e Google Acadêmico, utilizando os descritores: diabetes mellitus, tecnologia do DNA recombinante e tratamento de diabetes mellitus com DNA recombinante. Restringiu-se a busca a partir do ano 2000, selecionando 4 textos científicos. Resultados: A insulina é utilizada no tratamento da diabetes, tornando necessário a ampliação da produção, simultaneamente que diminuam a rejeição e possuam baixo custo. A princípio utilizavam-se insulina animal, porém haviam suprimentos limitados, comparado à demanda. Posteriormente, fabricaram a insulina humana semissintética, onde purificavam a insulina animal, assim, amenizando as reações imunes. Com a criação da tecnologia do DNA recombinante, foram produzidas as insulinas humanas recombinantes, elevando os benefícios devido sua pureza e melhor qualidade. Subsequente desenvolveram análogos da insulina que possui maior eficiência, pois, reduz casos de hipoglicemia, contudo, a insulina recombinante até então, é utilizada devido ao custo-benefício. Conclusão: Constata-se que a criação dessa biotecnologia foi um marco para o tratamento da diabetes, visto que possibilitou a fabricação de insulina com DNA humano em larga escala, com valor acessível e redução de rejeições, porém é mostrado obstáculos nesse tratamento, tendo potencial na combinação dessa tecnologia a outras, gerando inovações no tratamento da DM e de outras doenças.
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Hirschberg, M., A. Manoutchei, B. Klemens und B. Hofferberth. „CEREBRAL THROMBOLYSIS WITH INTRAVENOUSLY ADMINISTERED RECOMBINANT LOW- MOLECULAR-WEIGHT-UROKINASE AND RECOMBINANT PRO-UROKINASE IN A DOG MODEL“. In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643571.
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There is increasing evidence that recombinant prourokinase (rec-pro-UK) is a proenzyme which in vivo systems may induce activation of the fibrinolytic system with a better thrombus selectivity than that obtained with active urokinase.In order to study the effects of rec-pro-UK and low-molecular-weight-urokinase (LMW-UK) on acute stroke, a thrombus was induced in the middle cerebral artery (MCA) of anesthetized mongrel dogs (n=12). Occlusion of the vessel was confirmed by angiography. Following a 1 hour period of MCA occlusion, in six animals LMW-UK was administered intravenously at a dose of 4000 lU/kg/min. Angiographically confirmed thrombolysis occurred after 30 minutes. Thrombolysis by LMW-UK was accompanied by bleeding from all surgical wounds and consumption of plasminogen, alpha-2-antiplasmin and fibrinogen. Rec-pro-UK was administered to six other dogs in a LMW-UK-equivalent dosis. Thrombolysis was achieved after 30 minutes in all six cases without inducing a systemic lytic state. Neither in the LMW-UK-group nor in the group treated with rec-pro-UK intracerebral bleeding complications were observed on post mortem examination.Our findings indicate that intravenous administration of rec-pro-UK - because of the lack of systemic side-effects - may be a safe way of rapid thrombolysis of occluded cerebral arteries in acute stroke.
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Duarte, Amanda Alves, Renata Andrade De Oliveira, Eliane Pereira Cipolatti, Evelin Andrade Manoel und Martina Costa Cerqueira. „COMPARAÇÃO DO DESEMPENHO DE LIPASE COMERCIAL E RECOMBINANTE DE CANDIDA ANTARCTICA FRAÇÃO B EM PARTÍCULAS DE PMMA“. In I Congresso de Engenharia de Biotecnologia. Revista Multidisciplinar de Educação e Meio Ambiente, 2021. http://dx.doi.org/10.51189/rema/1356.
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Introdução: As lipases possuem alto poder catalítico, realizando diversos tipos de reações e por isso são conhecidas como excelentes alternativas nas indústrias têxtil, farmacêutica, alimentícia, biocombustíveis, entre outras. O processo de imobilização pode promover o aumento e a manutenção da estabilidade das enzimas e possibilitar o reuso desses biocatalisadores, diminuindo os custos do processo. Desta forma, têm se estudado cada vez mais o aperfeiçoamento dos processos de imobilização. A importância industrial da aplicação de lipases imobilizadas é notável, e o desenvolvimento de novos biocatalisadores poderá influenciar beneficamente no desenvolvimento e avanço da biocatálise. Objetivo: Comparar o desempenho de duas lipases: a lipase B de Candida antarctica comercial (CalB) e a sua análoga recombinante obtida por cultivo submerso (LipB) em partículas casca-núcleo de poli(metacrilato de metila) (PMMA/PMMA) em reações de hidrólise e esterificação. Material e Métodos: O suporte PMMA/PMMA foi inicialmente tratado com etanol e água destilada. Após, foi adicionado volume correspondente de uma solução de CalB ou LipB em solução tampão fosfato de sódio (5 mM, pH 7). A atividade do sobrenadante foi monitorada utilizando p-nitrofenil-laurato (p-NFL). A atividade hidrolítica foi medida no espectro utilizando como substrato p-NFL, com agitação suave e banho a 30ºC. A atividade de esterificação foi avaliada em ácido oleico e etanol (1:1) a 45ºC. Resultados: As enzimas foram imobilizadas com elevada porcentagem de imobilização, 95,92% para CalB e 96,2% para LipB em 4h. Na atividade hidrolítica os resultados foram CalB 53,84 e LipB 49,89 U/g biocat. Atividade de esterificação obtivemos para a CalB 519,61 e LipB 392,12 U/g biocat. Conclusão: O polímero núcleo-casca, poderá ser usado com sucesso para imobilização. O biocatalisador home-made em desenvolvimento apresentou muitas vantagens e tem seus destaque tanto para atividade enzimática quanto para aplicação em reações de esterificação comparada a CalB que é uma enzima comercial. Embora o trabalho ainda se encontre em desenvolvimento, os resultados são promissores e o biocatalisador obtido poderá ser usado em diversas áreas industriais.
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Camargo, Luiza Helena Angarten Ferraz de, Thalita Barcelos Silva, Lysandra De Queiroz Cunha Barradas, Dhullya Eduarda Resende Santos und Hanstter Hallison Alves Rezende. „O USO DA BIOTECNOLOGIA NO CONTEXTO DOS ALIMENTOS FUNCIONAIS“. In I Congresso de Engenharia de Biotecnologia. Revista Multidisciplinar de Educação e Meio Ambiente, 2021. http://dx.doi.org/10.51189/rema/1368.
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Introdução: A biotecnologia é resultado da combinação de sistemas biológicos e da tecnologia. Na indústria alimentícia, pode ser utilizada para otimizar e aperfeiçoar propriedades dos alimentos que podem ser benéficas para a saúde humana, como no caso dos alimentos funcionais, que além de serem capazes de oferecer grandes benefícios a saúde, são muito nutritivos. São exemplos destes a soja por conter isoflavonas e peixes que possuem ácidos graxos ômega 3. Objetivo: Realizar uma revisão bibliográfica sobre a importância da biotecnologia na procura por alimentos funcionais afim de favorecer a saúde humana. Material e métodos: Trata-se de uma revisão de literatura realizada a partir da seleção e análise de 4 artigos, publicados entre os anos de 2016 a 2021, sendo coletados nas plataformas de pesquisa Google Acadêmico e Periódicos CAPEs encontrados utilizando os descritores: biotecnologia, indústria e alimentos funcionais[HH1] . Resultados: Existe uma clara relação entre os alimentos consumidos e a saúde. Atualmente, os consumidores apresentam uma maior preocupação quanto ao consumo de alimentos que contribuam beneficamente para a saúde. Os alimentos funcionais possuem substâncias que atuam ativando processos metabólicos, que contribuem para uma melhoria do sistema imunológico. As propriedades deste tipo de alimento podem prover de constituintes naturais ou então de compostos adicionados artificialmente (como fibras, minerais e proteínas modificadas), afim de se obter um melhoramento nutricional dos mesmos. A biotecnologia utiliza de técnicas como a mutagênese, conjugação, genética recombinante e os bioprocessos fermentativos, para se obter componentes que possam acrescentar a formulação natural do alimento com o propósito de promover uma vida alimentar saudável do consumidor, prevenindo o aparecimento precoce de alterações patológicas e reduzindo o risco de doenças crônicas degenerativas e doenças crônicas não transmissíveis (DCNT), como diabetes e hipertensão. Conclusão: A biotecnologia aplicada no desenvolvimento dos alimentos funcionais traz cada vez mais benefícios para a saúde da população em geral, prevenindo e auxiliando no tratamento de diversas doenças, como a diabetes tipo 2, doenças cardiovasculares e também em casos de degenerações musculares.
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Bridi, Vanessa, Gabriela Alves Carvalho Duarte, Dhullya Eduarda Resende Santos und Hanstter Hallison Alves Rezende. „BIOFÁRMACOS: DESAFIOS ENFRENTADOS NA PRODUÇÃO NACIONAL“. In I Congresso de Engenharia de Biotecnologia. Revista Multidisciplinar de Educação e Meio Ambiente, 2021. http://dx.doi.org/10.51189/rema/1376.
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Introdução: Os biofármacos, também conhecidos como medicamentos biológicos, são fármacos desenvolvidos por meio de processos biotecnológicos, onde se utiliza a tecnologia do DNA recombinante em sistemas vivos como bactérias, vírus, leveduras e principalmente células de mamíferos. Com o crescente desenvolvimento tecnológico, cada vez mais a indústria farmacêutica vem se inovando neste âmbito, afim de combater inúmeras doenças que não são sensíveis a terapias tradicionais, como por exemplo, o câncer, artrite reumatoide, diabetes, dentre outras. Infelizmente, acompanhado do avanço biotecnológico vem alguns desafios importantes que atrapalham a produção em larga escala em nosso país. Objetivo: Descrever os principais desafios encontrados na produção de biofármacos no Brasil no âmbito público. Materiais e métodos: Trata-se de um trabalho de revisão bibliográfica, onde foram coletados 10 artigos, dos quais 6 foram selecionados para síntese deste trabalho. As bases de dados de procura foram: PubMed, SciELO e Google Acadêmico, utilizando os descritores: biofármacos e medicamentos biológicos, publicados na série temporal 2016-2020. Resultados: A produção de um biofármaco acompanha uma série de etapas que vão desde a descoberta de novas moléculas até a validação e caracterização de todo o processo para que se tenha um produto final. Entre essas etapas é necessária uma gama de processos industriais que necessitam de profissionais altamente capacitados, equipamentos sofisticados e investimentos altos para cada etapa de produção, que dura em média 10 a 15 anos ao todo. No Brasil, o principal desafio enfrentado é a falta de investimentos desde a pesquisa básica, tornando inviável a produção destes medicamentos no setor público, tendo a necessidade de importações, elevando os preços desses medicamentos no mercado e comprometendo a saúde populacional, principalmente das pessoas mais carentes que não possuem recursos para adquirir estes fármacos. Conclusão: Conclui-se que com o aumento da expectativa de vida, essas doenças acompanham a população corriqueiramente e que é indispensável a criação destes medicamentos para garantir uma melhor qualidade de vida. Sugere-se que sejam criados investimentos em indústrias e instituições nacionais para a produção destes fármacos e que sejam criadas novas estratégias para atender as demandas, como por exemplo a produção em larga escala de fármacos biossimilares.
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Gonçalves, Thalita Carvalho de, Dayane Silva Cavalcante, Martina Cerqueira Pinto, Eliane Pereira Cipolatti und Evelin Andrade Manoel. „APLICAÇÕES INDUSTRIAIS DE LIPASES IMOBILIZADAS: UM ESTUDO QUANTITATIVO“. In I Congresso de Engenharia de Biotecnologia. Revista Multidisciplinar de Educação e Meio Ambiente, 2021. http://dx.doi.org/10.51189/rema/1392.
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Introdução: As enzimas são catalisadores naturais de grande interesse industrial, pois, atuam sob condições operacionais mais brandas e apresentam alta atividade, seletividade e especificidade, possibilitando a geração de produtos mais puros e livres de contaminantes. A utilização de enzimas, como as lipases, têm aumentado nos últimos anos, sobretudo em áreas como alimentos, tecidos, fármacos, de cosméticos, de biocombustíveis, dentre outras, uma vez que são capazes de catalisar reações de hidrólise, esterificação e transesterificação. Além disso, com os avanços da tecnologia do DNA recombinante, é possível produzi-las em larga escala, tornando o processo ainda mais viável. A exploração do desenvolvimento de processos sustentáveis e economicamente viáveis tem despertado o interesse por tecnologias como a imobilização enzimática. A busca por processos de imobilização vem crescendo durante os últimos anos, principalmente devido aos avanços biotecnológicos e ao surgimento de novos materiais e técnicas de imobilização. Objetivo: Apresentar dados estatísticos da utilização de lipases imobilizadas nos últimos 10 anos, relacionando com o suporte utilizado e tipo de imobilização. Material e métodos: Os resultados estão sendo obtidos na plataforma PubMed e Scopus , através das buscas pelas palavras-chave lipase AND Immob* AND support. As publicações foram selecionadas segundo o seguinte critério: data de publicação de 2011 a 2021 e apenas publicações em inglês. Os seguintes parâmetros serão avaliados: frequência das técnicas e suportes utilizados nos últimos anos e aplicações das lipases. Os dados estão sendo cuidadosamente avaliados utilizando o programa Excel e serão organizados através do programa VosViewer para uma melhor visualização. Resultados e discussão: O trabalho está em desenvolvimento, mas resultados prévios mostram que, do total de publicações analisadas, as lipases imobilizadas se destacam nas áreas de bioquímica, genética e biologia molecular, seguida por medicina, química, engenharia química e agricultura. A técnica de imobilização por adsorção física é a mais utilizada (75% do total). Há uma tendência no surgimento de novos materiais e novas técnicas de imobilização, como intercruzamentos e nanoflowers. Conclusão: Os dados obtidos mostram que nos últimos 10 anos houve um aumento considerável pela busca do processo de imobilização. Estudos como este podem colaborar com o desenvolvimento da área e avanço da biocatálise